Research Of The Effect On Apoptosis And Secretion Of Granular Cell In Patients Suffered From Endomeriosis With QUYUJIEDU Therapy

Objective:EM-related infertility patients of blood stasis and toxin accumulation syndrome were treated with quyujiedu granule combined with IVF-ET. The efficacy and the effect on apoptosis and secretion of granular cells were observed and the potential mechanism was explored.Method:Clinical research: 82 cases of endometriosis(EM) related infertility patients were randomly divided into treatment group and control group with 42 cases and 40 cases respectively before IVF. The treatment group was given Quyujiedu granule orally for 3 months before IVF-ET. The control group was directly given IVF-ET. The symptom scores, pregnancy rate, content of E₂, P in serum and follicular fluid ,apoptosis rate of granule cell and the expression of Bcl-2 protein and Bax protein were measured. The E₂ and P secretion in granule cell after cultured for 48h in vitro was observed to learn about the secreting function of granule cell.Experiment 1: 40 EM rats were randomly divided into experiment group and control group with 20 in each group. Another 20 rats were processed by only stretching the uterus as pseudo-operation group. Quyujiedu granule was given by gastric injection in experiment group daily for 3 weeks and saline was given in the other 2 groups in the same way. All the rats received peritoneal injection of 50IU of HMG and HCG (100IU/100g) 24 hours and 48 hours after the final gastric injection respectively. 14-16 hours later, the rats were killed and the occytes and granular cells were collected in order to observe the number and rate of quality ovum. The apoptosis rate, Bax protein and Bcl-2 protein expression in the granular cells were measured using PI.Experiment 2: To dye the granule cells in nutrient solution by 0.4% trypan blue, take count of the live cells. When the livability of cells was higher than 90%, the suspl contains granule cells was adjusted to 3×10⁶ /ml monoplast with M199 nutrient solution (contains 20% calf serum, 100 U /ml penicillium and 100 mg/ml streptomycin) standby. Granule cells suspl was randomly implanted into culture dish which was imbrued by 0.05% L-polylysine beforehand by 3×10⁶ cells each hole, and then was precultured in 37℃homoiothermal couveuse contained 5%CO₂ and 95% air for 24h. To observe the growth of cells by inverted microscope. Exchange the solution with M199 for adherent cells and then culture for another 24h, 48h, afterwards, collect nutrient solution and cryopreserve it in -20℃for measure E2andP.Result:1.The symptoms was significantly improved and the pregnancy rate was elevated in the treatment group. Accordingly, the positive rate of EMAb and the CA₁₂₅ level decreased; the E₂ and P in patients serum and follicular fluid increased. Quyujiedu granule taken by treatment group could reduce apoptosis significantly rate of granule cell and improve the rate of fecundation, high quality ovum and embryo by depressing the expression of Bax protein , elevating the expression of Bcl-2 protein and then improving the ratio of Bcl-2 to Bax compared to control group (P＜0.05) .2.The amount and quality of high quality ovum in number and rate of treatment group was statistically higher than that of control group (P＜0.05). The apoptosis rate and Bax protein expression in granular cells of treatment group was significantly lower than control group(P＜0.05). Nevertheless, the Bcl-2 protein expression and Bcl-2/Bax ratio was higher in treatment group(P＜0.05).3.The E₂ and P of granule cells in after incubating 48h and 72h in vitro (＜0.05) . Conclusion: Quyujiedu granule could improve the symptoms and raise the pregnancyrate significantly. This may attribute to two main mechanism hereinafter: to reduce apoptosis rate of granule cell and improve the rate of fecundation ,high quality ovum and embryo by depressing the expression of Bax protein , elevating the expression of Bcl-2 protein and then improving the ratio of Bcl-2 to Bax; to advance secretion of granule cell in order to meliorate the state of incretion. Quyujiedu granule is an effective prescription for EM of blood stasis and toxin accumulation syndrome.